Lipid bilayers are thin polar membranes formed from two layers of lipid molecules. Lipid bilayers are found in cell membranes of most living organisms and are usually composed of phospholipids. They are impermeable to most hydrophilic molecules and ions, and enable cells to regulate their salt concentrations and pH by pumping ions across the lipid bilayer using transmembrane proteins known as ion pumps. Lipid bilayers, or more generally bilayers of amphipathic molecules, also serve as excellent platforms for a range of experimental studies. Holden et al, J. Am. Chem. Soc. 2007, 129, 8650-8655 disclose the formation of functional bionetworks of aqueous droplets comprising lipid bilayers provided between droplets. Such networks can act as light sensors, batteries and electrical components by incorporating pumps, channels and pores into the bilayers. Sackmann, Science, New Series, Vol 271, No. 5245 (Jan. 5, 1996), pp. 43-48 provides a review of the scientific and practical applications of supported lipid-protein bilayers including their use in electrooptical biosensors. Jung et al, J. Am. Chem. Soc., 2009, 131 (3), 1006-1014 have developed optical assays for the detection of protein ligand binding on supported bilayers. The provision of ion channels in highly resistive amphipathic lipid bilayers for the detection of DNA and other analytes is well documented, see for example Bayley et al, Nature, Vol 413, September 2001. Aqueous solutions are provided on either side of the lipid bilayer and ion flow through the nanopore takes place under a potential gradient. DNA is caused to translocate the ion channel and the change in ion flow during translocation of DNA through the channel is measured. Due to the high resistance of the lipid bilayer, ion flow takes place exclusively through the ion channel. The lipid bilayer may be suspended across an aperture of a substrate and formed by methods well known in the art such as patch clamping or painting.
WO2009/077734 discloses a plurality of individually addressable lipid bilayers formed across an array of microwell apertures, each microwell containing an electrode and an aqueous medium in contact with the lipid bilayer.
WO2009/012552 discloses a bilayer of amphipathic lipid molecules formed between two droplets comprising a layer of amphipathic molecules containing a hydrophilic medium, the droplets being provided in a hydrophobic medium. Ion flow across the lipid bilayer is measured with electrodes provided within the hydrophilic interior of each droplet.
An amphipathic molecule may be considered as comprising a polar hydrophilic region attached to a non-polar hydrophobic region. A bilayer may be formed from two monolayers of amphiphilic molecules, wherein in aqueous solution, the polar groups face towards the hydrophilic media on either side of the bilayer and the hydrophobic groups face inwards.
WO2009/024775 discloses a method for producing a droplet interface bilayer (DIB) wherein droplets are prepared by contacting an oil/lipid solution with an aqueous solution and the resulting droplets are brought into contact with an aqueous agarose gel support layer.
Phospholipids such as 1,2-diphytanoyl-sn-glycero-3-phosphatidylcoline (DPhPC) are routinely used to form lipid bilayers. However drawbacks that are sometimes associated with lipid bilayers include that they are not particularly robust and are prone to rupture, for example by digestion by enzymes, and are not able to withstand large potential differences.
U.S. Pat. No. 6,723,814 discloses a planar membrane formed from amphiphilic copolymers having hydrophilic and hydrophobic segments. The copolymer may be an ABA triblock having methyloxazoline hydrophilic segments and a dimethylsiloxane hydrophobic core (PMOXA-PDMS-PMOXA). Membranes formed from this triblock are able to withstand higher potential differences than lipid membranes (Table 1 of U.S. Pat. No. 6,723,814).
U.S. Pat. No. 6,916,488 describes the preparation of vesicles made from PMOXA-PDMS-PMOXA in a hydrophilic medium (type ABA). The structure of an amphipathic ABA triblock vesicle (a droplet in a hydrophilic medium having a hydrophilic interior) may be conceptualised as a monolayer of triblock polymer in which the polymer molecules have a linear configuration in which the two hydrophilic ‘A’ segments face the respective hydrophilic solutions on either side of the vesicle wall. Such a configuration, which is shown in FIG. 1 of U.S. Pat. No. 6,916,488, would not however seem suitable for stabilising aqueous droplets in oil. Such ABA molecules do not therefore seem to be a viable alternative to the lipids described in WO2009/024775, for producing a droplet interface layer from a water-in-oil system.
There is thus an ongoing need to provide alternative methods for producing interface membranes that provide improved stability compared to conventional lipid bilayers.